Figure 1.

3D-printed hanging drop dripper (3D-phd) for tumor spheroids study. (A) The workflow of 3D-phd for studying tumor spheroid generation, drug-induced cell death, and metastasis in extracellular matrix gel. The device has been printed and directly used for cell spheroid generation on a 96/384 well culture plate. Closeup artwork shows a detailed structure of an individual spheroid culture site with a cell aggregate. Four different assays have been developed based on 3D-phd platform: (i) Specific components for drug screening can be added through device, and cell toxicity imaging was directly acquired by dripping down treated cell spheroid with specific staining; (ii) 3D metastasis of cell spheroid on/in matrix could be performed on a matrix gel-coated plate or dropping off with cell-matrix gel solution directly; (iii) The 3D tumor spheroid based transendothelial migration analysis; (iv) Heterotypical spheroids interaction by using the double nozzles 3D-phd. Photograph of 3D-phd mounted on a 96 well plate shown in (B), and the closeup of the hanging drops on the device has been shown in (C). The standard pipetting operation for medium changing or dropping down is displayed in (D). (E) A 3D-phd cultured MCF-7 cell spheroid was stained by phalloidin-rhodamine (red) and Hoechst-33342 (blue). The staining was performed after 24 h culture. Scale bar is 200 µm. (F) The photograph of the hanging drops on double nozzles device (left) mounted on a 96-well U-shape low attachment plate. The time-lapse fluorescence images of the fusion dynamics of two different spheroids. The spheroids have been directly dropped down into the well, MCF-7 (CellTracker Red labeled) and MDA-MB231 (CellTracker Green labeled) tumor, respectively. Scale bar is 10 mm in the left photograph and 200 μm in fluorescence micrograph.